Endophytic Fungi from Uruguayan Native Myrtaceae: Enzymes Production, Antimicrobial and Phytotoxic Activity

Author(s): Lina Bettucci, Susana Tiscornia

In recent years fungal endophytes occurring in native Myrtaceae from Uruguay have been studied. Fungal associations with the host, such as saprotrophs, latent plant pathogens or symbiotic, are related with enzymes and secondary metabolites production. Therefore, a main goal of this work was to evaluate the ability of endophytic fungi isolated from Myrcianthes cisplatensis, Myrrhinium atropurpureum and Eugenia uruguayensis to produce hydrolytic and oxidative extracellular enzymes and bioactive metabolites with antimicrobial activity or phytotoxic properties. Enzymes production were evaluated by plate assay and metabolites were extracted with organic solvents. For detecting phytotoxicity, fungal extract were injected on fresh leaves of E. uruguayensis. Antimicrobial activity was evaluated by antibiogram technique and growth inhibition assessed by halo. Cladosporium sphaerospermum, Xylaria acuta and Amphisphariaceae 143 that showed the highest enzymatic activity probably degrading plant debris when they arrive to soil having saprotrophic life style. The enthomopathogenic fungus Metarhizium anisopliae producing proteases and chitinases is active pathogen of several insect species. In addition, Nigrospora spherica and Xylaria sp. produced necrotic spots on Myrtaceae leaves evidencing phytotoxic activity. The antifungal activity showed by some endophytic species evidenced the ability to limit the development of microorganism populations. Lophiostoma sp. 246 was the most active strain against Staphyloccocus aureus and Candida albicans and Preussia. africana evidenced a good antimicrobial activity against Xanthomonas campestris. Native Myrtaceae seems to be a good source of fungal endophytes for producing enzymes related with decomposition process and a promising source of bioactive metabolites with antimicrobial and fitotoxigenic activities. In addition, the new strain of M. anisopliaes could be a promising bioinsecticide.

Secondary metabolites, endocellulases, pectinases, ligninases, proteases, chitinases

1. Aly AH, Debbab A, Kjer J, Proksch P (2010) Fungal endophytes from higher plants: A prolific source of phytochemicals and other bioactive natural products (Review). Fungal Diversity 41: 1-16
2. Baüer A, Kirby W, Sherris I, Turk M (1966) Antibiotic susceptibility testing by standarized single disk method. American Journal of Clinical Pathology 45: 493-496
3. Berestetskiy A (2008) A Review of Fungal Phytotoxins: from Basic Studies to Practical Use. Applied Biochemistry and Microbiology 44: 453-465
4. Bettucci L, Speranza M, Piaggio M, Siñeris F, Breccia JB (1998) Degradation of Eucalyptus globulus and Eucalyptus grandis by several white rot fungi isolated in South America. Material und Organismen 32: 227-233
5. Bettucci L, Simeto S, Alonso R, Lupo S (2004) Endophytic fungi of twigs and leaves from three wild species of Myrtaceae. Sydowia 56: 8-23
6. Boddy L, Griffith GS (1989) Role of endophytes and latent invasion of the development of decay communities in sapwood of angiospermous trees. Sydowia 41: 41-73
7. Breccia JD, Siñeriz F, Bettucci L, Piaggio M (1997) Degradation of sugar cane bagasse by several white-rot fungi Acta Biotechnologica 17:177–184
8. Brussa CA, Grela IA (2007) Flora arbórea del Uruguay. COFUSA. Uruguay
9. Chul-Kang S, Park S, Gyu-Lee D (1999) Purification and characterzation of a novel chitinase from the entomopathogenic fungus, Metarhizium anisopliae. Journal of Invertebrate Pathology 73: 203-215
10. Davis EC, Franklin JB, Shaw AJ, Vilgalys R (2003) Endophytic Xylaria (Xylariaceae) among liverworts and angiosperms: phylogenetics, distribution, and symbiosis. American Journal of Botany 90: 1661-1667
11. Dreyfuss MM, Chapela IH (1994)Potencial of fungi in the discovery of novel, low-molecular weight pharmaceuticals, p. 49-80. In V. P. Gullo (ed.) The discovery of natural products with therapeutic potencial. Butter-worth-Heinemann, London, UK
12. Eggins HOW, Pugh GJF (1962) Isolation of cellulase decomposing fungi from soil. Nature 193: 94-95
13. Fenice M, Federici F, Giavannazzi Sermanni G, D’Annibale A (2005) Submerged and solid-state production of lacasse and Mn-peroxidase by Panus tigrinus on olive mill wastewater-based media. Journal of biotechnology 100: 77-85
14. Hormazabal E, Piontelli E (2009) Endophytic fungi from Chilean native gymnosperms: antimicrobial activity against human and phytopathogenic fungi. World J Microbiol Biotechnol 25:813-819. DOI 10.1007/s11274-008-9953-6
15. Lv Y, Zhang F, Chen J, Cui J, Xing Y, Li X, Guo S (2010) Diversity and antimicrobial activity of endophytic fungi associated with the alpine plant Saussurea involucrate. Biological and Pharmacological Bulletin 233:1300-1306
16. Machado KMG, Matheus DR, Bononi VLR (2005) Lignolytic enzyme production and Remazol Brillant Blue R decolorization by tropical brazilian basidiomycetes fungi. Brazilian Journal of Microbiology 36: 246-252
17. Mier T, Rosas-López B, Castellanos-Moguel J, García-Gutiérrez K, Toriello C (2004) Efecto de la citrolina sobre la viabilidad y la producción de proteasas y quitinasas del hongo entomopatógeno Metarhizium anisopliae var. acridium. Revista Mexicana de Micología 19: 113-115
18. Oliveira K F, Malavolta L, Souza CS, Vicente EJ, Laluce C (2006) Pectinolytic activity secreted by yeasts isolated from fermented citrus molasses. Journal of Applied Microbiology 100: 633-640
19. Oses R, Valenzuela S, Freer J, Baeza J, Rodríguez J (2006) Evaluation of fungal endophytes for lignocellulolytic enzyme production and wood biodegradation. International Biodeterioration & Biodegradation 57: 129-135
20. Petrini O, Petrini LE, Rodrigues K (1995) Xylariaceous endophytes: An Exercise in Biodiversity. Fitopatol Bras. 20: 531-539
21. Pointing SB, Parungao MM, Hyde KD (2003) Production of wood decay enzymes, mass loss and lignin solubilization in wood by tropical Xylariaceae. Mycol. Res 107: 231-235
22. Promputtha I, Lumyong S, Dhanasekaran V, Mckenzie EHC, Hyde KD, Jeewon R ( 2007) A phylogenetic evaluation of whether endophytes become saprotrophs at host senescence. Microb Ecol 53: 579-590
23. Promputtha I, Hyde KD, McKenzie EHC, Peberdy JF, Lumyong S (2010) Can leaf degrading enzymes provide evidence that endophytic fungi becaming saprobes ? Fungal Diversity 41:89-99
24. Saikkonen K (2007) Trees can not escape endophyte infections. Fungal Biology Reviews 21: 67-74
25. Smedsgaard J (1997) Micro-scale extraction procedure for standarized screening of fungal metabolite production in cultures. J. Chromatogr. 760: 264-270
26. Sieber TN (2007) Endophytic fungi in forest trees: are they mutualists? Fungal Biology Reviews 21:75-89
27. Shirouzu T, Hirose D, Fukasawa Y, Tokumasu S (2009) Fungal succession associated with the decay of leaves of an evergreen oak Quercus myrsinaefolia. Fungal Divers. 34:87–109
28. Suryanarayanan TS, Thirunavukkarasu N, Govindarajulu MB, Gopalan V (2012) Fungal endophytes: an untapped source of biocatalysts. Fungal Diversity. DOI 10. 107/s13225-012-0168-7
29. Tiscornia S (2012) Comunidades de hongos endófitos de Myrtaceae neotropicales: evaluación de la producción de enzimas y metabolitos bioactivos. PhD Thesis. Universidad de la Republica. Uruguay
30. Tiscornia S, Ruiz R, Bettucci L (2012) Fungal endophytes from vegetative and reproductive tissues of Eugenia uruguayensis in Uruguay. Sydowia 64: 313-328
31. Varela H (1998) Producción de enzimas depilantes de pieles. PhD Thesis. Universidad de la República. Uruguay
32. Whalley AJS (1996) The xylariaceous way of life. Mycol. Res.100: 897-922